I Hormones of the Anterior Pituitary
The anterior pituitary (adenohypophysis) is the source of several major glycoprotein hormones including luteinizing hormone (lutropin, or "LH"), choriogonadotropin (or "CG"), follicle stimulating hormone (follitropin or "FSH"), and thyroid stimulating hormone (thyrotropin or "TSH"). The hormones of the anterior pituitary are reviewed by Norman, A. W. et al. (In: Hormones, Acad. Press, New York., (1987)). The hormones are highly conserved evolutionarily; the primary amino acid sequences of the LH, CG, and TSH hormones of rat and other animals are highly similar to those of humans (Strickland, T. W. et al., In: Luteinizing Hormone Action and Receptors, Ascoli, M. (Ed.), CRC Press, Boca Raton, Fla. (1985)).
Luteinizing hormone, follicle stimulating hormone, human choriogonadotropin (hCG) and thyroid stimulating hormone share many common characteristics, and have been considered to be members of a family of glycoprotein hormones. All contain approximately 200-250 amino acid residues, and are composed of a common .alpha. subunit (having a molecular weight of approximately 13-15 kDa) and a distinctive .beta. subunit (having a molecular weight of approximately 13-22 kDa). The .alpha. subunits of LH, FSH, and TSH are identical; the .alpha. subunit of CG has been reported to differ slightly from the others (Ganong, W. F. Review of Medical Physiology, 9th Ed., Lange Medical Pub., Los Altos, Calif., (1979)).
The hormones mediate their biological actions by binding to receptor molecules present on the surfaces of target cells. Interaction of the .alpha. subunit with the hormone specific .beta. subunits of the hormones are responsible for confering the binding specificity of the hormones. The hormones act by activating cellular adenylate cyclase to increase intracellular cAMP levels (de la Llose-Hermier et al., Acta Endocrinol, 11:399-406 (1988)).
Luteinizing hormone and follicle stimulating hormone are both gonadotropins (Ascoli, M. (Ed.) Luteinizing Hormone Action and Receptors, CRC Press, Boca Raton, Fla., (1985)). LH binds to a receptor expressed on the surfaces of Leydig (interstitial) cells (Ascoli, M., In: The Receptors, (Conn, P. M. (Ed.), vol. 2, pp 368 (1985)). In men, LH binding causes the Leydig cells to increase their synthesis of testosterone. In women, such binding causes the granulosa, theca, interstitial, and luteal cells to increase the concentrations of androgens, estrogens, and progestins, especially progesterone.
Follicle stimulating hormone regulates the development of gametes. In men, FSH binds to a receptor present on the surface of the Sertoli cells, and assists in the developmental process which results in the production of mature spermatozoa. In women, the hormone binds to receptors on the surface of the granulosa cells of the ovary. It is believed to act in concert with estrogen and LH to stimulate follicle development. Reflecting its role in oocyte development, FSH is maximally expressed at the time of ovulation in the female reproductive cycle. For this reason, an assay for FSH can be used to detect and predict the occurrence of ovulation. FSH also acts to stimulate the expression of LH/CG receptors by granulosa cells.
Choriogonadotropin is a gonadotropin produced by the trophoblastic cells of the placenta. It acts to stimulate the growth and development of the corpus luteum in the ovary by stimulating the production of progesterone. Choriogonadotropin has a role in preparing the maternal metabolism for the pregnancy. CG expression increases rapidly after conception, and as such, can be used as an assay for pregnancy. Administration of either LH, FSH, or CG can induce ovulation in a female. The hormones may be used in the treatment of infertility.
The principal action of TSH is to stimulate thyroid secretion and growth. TSH binds to a receptor molecule which is expressed on the surface of thyroid cells. Monoclonal antibodies have been developed which are able to bind to the TSH receptor. Individuals suffering from Graves Disease produce autoantibodies which are capable of binding to the TSH receptor molecule. In contrast to the anti-TSH receptor monoclonal antibodies, binding by the autoantibodies mimics TSH, and therefore acts as a stimulator of thyroid activity (Furmaniak, J. et al., Acta Endocrinol, (Suppl) 281: 157-165 (1987)). The clinical symptoms of Graves Disease are marked by hyperthyroidism.
II Receptors of the Glycoprotein Hormones of the Anterior Pituitary (including hCG)
Studies have revealed that luteinizing hormone and choriogonadotropin share the same cellular receptor molecule (Ascoli, M. (Ed.) Luteinizing Hormone Action and Receptors, CRC Press, Boca Raton, Fla., (1985)). The use of chemical and photoaffinity cross-linking agents has enabled researchers to study the hormone binding site of the receptor (Ji, I. et al., Proc. Natl. Acad. Sci. (U.S.A.) 77:7167 (1980); Rebois, R. V. et al., Proc. Natl. Acad. Sci. (U.S.A.) 78:2086 (1981); Metsikko, M. K. et al., Biochem J. 208:309 (1982)). These studies have not, however, led to the elucidation of the structure of the receptor molecule. On the basis of such studies, several research groups have concluded that the receptor is a single polypeptide of approximately 70-105 kDa (Ascoli, M. et al., J. Biol. Chem. 261:3807 (1986); Rebois, R. V. et al., Proc. Natl. Acad. Sci. (U.S.A.) 78:2086 (1981); Kellokumpu, S. et al., Endocrinol. 116:707 (1985)). Similar studies, however, have led other researchers to conclude that the receptor was composed of several polypeptide subunits (Ji, I. et al., Proc. Natl. Acad. Sci. (U.S.A.) 77:7167 (1980); Ji, I. et al., Proc. Nat. Acad. Sci. (U.S.A.) 78:5465 (1981); Hwang, J. et al., J. Biol. Chem. 259:1978 (1984); Hwang, J. et al., Proc. Natl. Acad. Sci. (U.S.A.) 81:4667 (1984)). The disparate conclusions reached by these researchers have not been reconcilable.
The purification of the LH/CG receptor has been reported by several research groups (Dufau, M. L. et al., J. Biol. Chem. 250:4822 (1975); Kusuda, S. et al., J. Biol. Chem. 261:6161 (1986); Bruch, R. C. et al., J. Biol. Chem. 261:9450 (1986); Minegishi, T. et al., J. Biol. Chem. 262:17138 (1987); Wimalasena, J. et al., J. Biol. Chem. 260:10689 (1985); Keinanan, K. P. et al., J. Biol. Chem. 262:7920 (1987); Dattatreyamurty, B. et al., J. Biol. Chem. 258:3140 (1983)). The reported characteristics of the purified protein have, however, been so disparate that they have failed to permit a conclusion to be reached regarding the nature of the LH/RH receptor, or the number of subunits which it contains.
The FSH receptor has also not been well characterized. Using photoaffinity techniques, researchers have concluded that the FSH receptor is composed of three subunits (Shih, J. et al., J. Biol. Chem. 260:12822 (1985); Shih, J. et al., J. Biol. Chem. 260:12828 (1985); Shih, J. et al., J. Biol. Chem. 260:14020 (1985); Smith, R. A. et al., J. Biol. Chem. 260:14297 (1985); Smith, R. A. et al., J. Biol. Chem. 260:14297 (1985)). The TSH receptor has been reported to be an approximately 300 kDa protein, which is cleaved to form at least two 70 kDa proteins. The 70 kDa proteins can themselves be cleaved to form a 50 kDa and a 20 kDa protein (Chan, J. et al., Acta Endocrinol. (Suppl.) 281:166 (1987); Smith, R. A. et al., Endocrinol. Rev. 9:88 (1988)).
In summary, the glycoprotein hormones of the anterior pituitary, as well as choriogonadotropin made by the placenta, have been found to mediate their biological effects via an interaction with a cellular receptor molecule present on the surface of target cells. Despite vigorous efforts, the nature and structure of these receptor molecules has not been resolved.
The hormone receptor molecules may be used both for diagnostic and therapeutic purposes. The receptor molecules may also be used to design synthetic hormones or hormone antagonists. Thus, an ability to produce purified hormone receptor molecules would be highly desirable.